Deactivants for dust mite allergens

ABSTRACT

Der-f and/or Der-p dust mite allergens are deactivated by a liquid composition comprising one or more of the following deactivants: i) cedarwood oil, ii) hexadecyltrimethylammonium chloride, iii) aluminium chlorohydrate, iv) 1-propoxy-propanol-2, v) polyquaternium-10 vi) silica gel, vii) propylene glycol alginate, viii) ammonium sulphate, ix) hinokitiol, x) L-ascorbic acid, xi) immobilised tannic acid, xii) chlorohexidine, xiii) maleic anhydride, xiv) hinoki oil, xv) a composite of AgCl and TiO 2 , xvi) diazolidinyl urea, xviii) a compound of formula I 
                         
xix) the compound of formula II
 
                         
xx) a polymeric dialdehyde containing two or more of a recurring unit of the formula III
 
                         
where n=2 to 200, xxi) urea, xxii) cyclodextrin, xxiii) hydrogenated hop oil, xxiv) polyvinylpyrrolidone, xxv) N-methylpyrrolidone, xxvi) the sodium salt of anthraquinone, xxvii) potassium thioglycolate, and xxviii) glutaraldehyde. Deactivants (i) to (xx) are effective against allergens derived from both species. Deactivants (xxi) to (xxvi) are effective against only Der-f allergens. Deactivants (xxvii) and (xxviii) are effective against only Der-p allergens.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. application Ser. No.09/509,308, filed May 25, 2000 now U.S. Pat No. 6,800,247, which is theU.S. National Phase filing of PCT Application No. PCT/GB98/02863 filedSep. 22, 1998, which claims priority to British Applications No.9720275.8 and 9720298.0 both filed Sep. 25, 1997. The disclosures of allthree prior applications are considered part of (and are incorporated byreference in) the disclosure of this application.

BACKGROUND OF THE INVENTION

It has been known for a long time that house dust can trigger allergenicreactions in humans, such as asthma and rhinitis. It was reported, asearly as 1928, that it was the dust mites in the dust that were theprimary source of the allergenic response but it was only in the 1960'sthat researchers appreciated its significance.

It is believed that the faeces of two particular house dust mite,species, Dermatophagoides forinae (known as Der-f) and Dermatophagoidespteronyssinus (known as Der-p) trigger the immune responses of the body,thereby giving rise to well known allergenic symptoms.

A review of this is given in Experimental and Applied Acarology, 10(1991) p. 167-186 in an article entitled “House dust-mite allergen”: Areview by L. G. Arlian.

Both the Der-f and Der-p species are found throughout the world. In someareas, Der-f will be the sole Dermatophagoides species. In other areasDer-p will be the sole species. In still other areas, the two speciesare both present through, generally, one or the other will predominate.

One way to overcome these allergenic response has been to vacuumsurfaces, such as carpets, that contain the dust mites and their faecesthoroughly and often, but that is both time consuming (i.e. has to beregularly done if one wants to make an allergenic free environment) andis very dependant on the efficiency of vacuum cleaner and filter bagused e.g. micron filter bag or 2-layer vacuum bags.

An alternative method of creating an allergen-free environment has beento denature the allergen, for example as described in U.S. Pat. No.4,806,526. The only effective method however of which we are aware is toapply tannic acid to the allergen. However, tannic acid can causestaining, and this is a particularly acute problem for light colouredcarpets (e.g. white and light-beige carpets) and other textile surfacesas tannic acid leaves a deep brown stain.

Therefore, we have been looking for allergenic denaturants which willnot stain susceptible surfaces such as carpets and still deactivate theallergen

We have discovered a number of allergen deactivants which are effectiveagainst both the Der-f and the Der-p species. Quite surprisingly, wehave discovered that some of these deactivants are specific to the typeof dust mite allergen being treated for example an effective Der-fallergen deactivants will not automatically work effectively as a Der-pallergen deactivant.

DISCLOSURE OF THE INVENTION

According to the invention there is provided a method for deactivatingallergens derived from the Der-f and/or Der-p dust mite species, whichcomprises contacting the allergen with a deactivating effective amountof one or more of deactivants (herein after defined as the deactivant).

The deactivants effective against one or both of Der-f allergens andDer-p allergens are:

-   -   i) cedarwood oil,    -   ii) hexadecyltrimethylammonium chloride,    -   iii) aluminium chlorohydrate,    -   iv) 1-propoxy-propanol-2,    -   v) polyquaternium-10    -   vi) silica gel,    -   vii) propylene glycol alginate,    -   viii) ammonium sulphate,    -   ix) hinokitiol,    -   x) L-ascorbic acid,    -   xi) “immobilised tannic acid”, (hereinafter defined)    -   xii) chlorohexidine,    -   xiii) maleic anhydride,    -   xiv) hinoki oil,    -   xv) a composite of AgCl and TiO₂,    -   xvi) diazolidinyl urea,    -   xvii) 6-isopropyl-m-cresol,    -   xviii) a compound of formula I

-   -   xix) the compound of formula II

-   -   xx) a polymeric dialdehyde containing two or more of a recurring        unit of the formula III

where n=2 to 200,

-   -   xxi) urea,    -   xxii) cyclodextrin,    -   xxiii) hydrogenated hop oil,    -   xxiv) polyvinylpyrrolidone,    -   xxv) N-methylpyrrolidone,    -   xxvi) the sodium salt of anthraquinone,    -   xxvii) potassium thioglycolate, and    -   xxviii) glutaraldehyde        Deactivants (i) through (xx) are effective against both Der-f        and Der-p allergens. Deactivants (xxi) through (xxvi) are        effective against Der-f allergens only. Deactivants (xxvii)        and (xxviii) are effective against Der-p allergens only.

A compound of formula I is commercially available as Aerosol OT.

The compound of formula II is commercially available as parsley camphor.

Hinoki oil is a mixture of thujan-3-one, 2-pinene,3,5,7,3′,4′-pentahydroflavanone and 1,3,3-trimethyl-2-norcamphanone.

Hydrogenated Hop Oil is the potassium salt of tetrahydroiso humulinicacid (also known as reduced isomerised hop extract).

Propylene glycol alginate is

Chlorohexadine is 1,1′-hexamethylenebis [5-(4-chlorophenyl)]-biguanide.

Hinokitol is β-thujaplicin, a compound of the formula

Germall II is diazolidinylurea.

Thymol is 6-isopropyl-m-cresol.

Cedarwood oil contains α- and β-cedrene (ca 80%), cedrol (3-14%) andcedrenol. Other sesquiterpenes and some monoterpenes are also present.

Polyquaternium-10 is a polymeric quaternary ammonium salt ofhydroxyethyl cellulose reacted with a trimethyl ammonium substitutedepoxide commercially available as Polymer JR-125.

Silica gel is also known as colloidal silica or silicic acid and iscommercially available as Kent.

“Immobilised tannic acid” is tannic acid on polyvinyl pyrrolidone beads.Immobilised Tannic Acid was prepared as follows: 100 mg of tannic acidwas dissolved in water; 50 mg of Polyclar 10 (ISP, Guildford Surrey)polyvinyl pyrrolidone beads were added and stirred for one hour; thebeads were filtered off the solution and washed with a few mls of icedwater until no colour was seen in the washings; they were then dried inthe oven at 50° C.

The composite of silver chloride and TiO₂ is made up of 20% wt/wt AgClon 80% TiO₂ 3-5 μm porous beads.

In compositions containing the deactivant, the deactivant is present inan amount of from 0.01% to 7%, preferably from 0.01% to 3%.

In methods for treating rugs and carpets to deactivate allergents, theamount of deactivant present is from about 16 gm to about 170 gm per 10square meters, preferably about 32 gm per 10 square meters.

Preferably the deactivant is selected from

-   -   xiv) hinoki oil,    -   xv) a composite of AgCl and TiO₂,    -   xvi) diazolidinyl urea    -   xvii) 6-isopropyl-m-cresol,    -   xii) chlorohexidine,    -   xiii) maleic anhydride,    -   xxvi) the sodium salt of anthraquinone and    -   xviii) a compound of formula I or II, defined above, and    -   xix) a compound of formula II, defined above.

Further according to the invention there is provided an aerosolcomposition containing.

-   -   i) cedarwood oil,    -   ii) hexadecyltrimethylammonium chloride,    -   iii) aluminium chlorohydrate,    -   iv) 1-propoxy-propanol-2,    -   v) polyquaternium-10    -   vi) silica gel,    -   vii) propylene glycol alginate,    -   viii) ammonium sulphate,    -   ix) hinokitiol,    -   x) L-ascorbic acid,    -   xi) “immobilised tannic acid”, (hereinafter defined)    -   xii) chlorohexidine,    -   xiii) maleic anhydride,    -   xiv) hinoki oil,    -   xv) a composite of AgCl and TiO₂,    -   xvi) diazolidinyl urea,    -   xvii) 6-isopropyl-m-cresol,    -   xviii) a compound of formula I

-   -   xix) the compound of formula II

-   -   xx) a polymeric dialdehyde containing two or more of a recurring        unit of the formula III

where n=2 to 200,

-   -   xxi) urea,    -   xxii) cyclodextrin,    -   xxiii) hydrogenated hop oil,    -   xxiv) polyvinylpyrrolidone,    -   xxv) N-methylpyrrolidone,    -   xxvi) the sodium salt of anthraquinone,    -   xxvii) potassium thioglycolate, and    -   xxviii) glutaraldehyde        b) a propellant, and        c) optionally, a solvent.

Preferably the amount of deactivant present in such a composition isfrom 0.01% to 7%, more preferably 0.01% to 3%,

Preferably the amount of propellant present in such a composition isfrom 4% to 50%, more preferably from 4% to 30%,

Preferably the amount of solvent present in such a composition is 0% to99.95%, more preferably 0% to 90%, and most preferably from 20% to 90%.

Preferably the deactivant in such aerosol composition is selected from

hinoki oil,

a composite of AgCl with TiO₂,

diazolidinyl urea,

6-isopropyl-m-cresol,

chlorohexidine,

maleic anhydride,

the sodium salt of anthraquinone, and

a compound of formula I or II defined above.

Preferably the propellant is selected from those commercially available,for example C₁₋₄ alkanes, chlorofluorocarbons and compressed gases suchas nitrogen, air and carbon dioxide.

Preferably the solvent is selected from C₁₋₆ alcohols (e.g. ethanol) orwater.

In addition, the compositions of this invention may also contain one ormore of the following:

-   -   a fragrance, preferably in an amount of 0% to 5%, more        preferably 0% to 2%;    -   an antimicrobial compound e.g. alkyldimethylbenzyl ammonium        saccharinate, preferably in an amount of 0.01% to 1%;    -   a surfactant, e.g. Dow Corning 193 Surfactant, preferably in an        amount of 0.01% to 1%;    -   a corrosion inhibitor, e.g. sodium nitrite, sodium benzoate,        triethanolamine and ammonium hydroxide, preferably in an amount        of 0.01% to 10%; and    -   a miticide, such as benzyl benzoate, pyrethroid pemethrin,        d-allethrin and optionally a synergist such as piperonyl        butoxide, preferably in an amount of 0.1% to 10%.

It has been found that deactivants of the invention have as effectiveallergen deactivating properties as tannic acid but without the drawbackof staining.

The invention will now be illustrated by the following Examples.

EXAMPLES

The test procedure in Examples 1 to 55 is as follows and is known as theELISA protocol.

The ELISA protocol for Der-f and Der-p has been developed as follows asa measure of denaturing property for denaturants.

ELISA Protocol 1

-   1. Dust is collected from Hoover™ vacuum cleaner bags and passed    through a series of sieves down to 63 microns.-   2. Clean petri dishes are labelled with the chemical to be tested    (on the base). Three replicates are used for each treatment.-   3. Filter paper is used to line each dish and 0.2 g of dust is added    to each dish onto the filter paper. The lid (or base, as dishes are    inverted) is replaced and the dish is shaken to disperse the dust    evenly over the filter paper.-   4. 2% aqueous solutions of deactivant were used except for the    silver chloride composite where 0.05% was used instead. Immobilised    tannic acid was used as a 1% dispersion. The hydrogenerated hop end    was used at the 2% level (in the form of a 10% solution).    Water-insoluble deactivants were emulsified with a sorbitone oleate    surfactant (i.e. Tween). Hinokitol was used at 0.5% not 2%.-   5. The dust is sprayed with the corresponding treatment, 2 sprays    are required for sufficient coverage (1 spray=1.5 ml).-   6. Leave uncovered at room temperature, in well aerated room, until    filter paper is dry. This can take up to 4 hours.-   7. Empty dust in epindorfs labelled according to treatment.-   8. Add 1 ml of 5% Bovine Serum Albumen Phosphate Butter Saline-Tween    BSA-PBS-T to each epindorf (5 times the weight of dust) (20 ml of    BSA-PBS-T=1 g of BSA in 20 ml of PBS-T).-   9. Leave overnight in a refrigerator.-   10. Centrifuge for 5 minutes at 13,000 rpm.-   11. Decant the supernatant into a new epindorf labelled according to    treatment.-   12. Centrifuge again for 5 minutes at 13,000 rpm.-   13. Make up dilutions of 1:10 and 1:100 by adding 100 μl of neat    solution to 900 μl of 1% BSA-PBS-T (1:10) This is repeated using 100    μl of 1:10 dilution and add to 900 μl of 1% BSA-PBS-T for 1:100    dilution.    ELISA Protocol 2 for Der-f and Der-p: Indoor Biotechnologies-   1. Prepare samples and dilutions as in protocol-   2. Prepare 500 ml of 50 mM carbonate/bicarbonate buffer by    dissolving 0.795 g Na₂CO₃ and 1.465 g NaHCO₃ in 500 ml of distilled    water. Check the pH is at 9.6. (This solution is kept in the    refrigerator in a conical flask).-   3. Monoclonal antibody (kept in the freezer) has to be added to the    buffer using the following method, (1 μg per well; 11 ml is needed)    applied to the ELISA plate    -   11 ml of carbonate/bicarbonate buffer is added to the dispensing        tray.    -   11 μl of Der-f1 or Der-p1 monoclonal antibody        (Stored in freezer, epindorf in use is in the refrigerator) is        added to the buffer. To ensure that all the antibody is removed        from the tip, wash out the pipette tip by sucking up and down I        the buffer solution, gently stirring to mix thoroughly.-   4. Pipette 100 μl of the antibody solution into each well of the    microtiter plate, cover with a plate sealer and leave overnight at    4° C.-   5. Empty the plate by quickly inverting it over the sink, then dry    by banging on a stack of paper towels.-   6. Add 200 μl of wash buffer to each well: PBS/0/05% tween (PBS-T).-   7. Repeat stages 5 and 6 once more (making a total of 2 washes).-   8. Make sure all the wells are dry, then add 100 μl of 1% BSA-PBS-T.    Replace the plate sealer and incubate for 1% at room temperature*.-   9. Repeat steps 5 to 7 (2 washes)-   10. *During the hour incubation period, prepare the allergen    standards at dilutions between 125 and 1 μg/ml Der f 1 or Der p1:    -   Add 25 μl of allergen standard (kept in the refrigerator in        polystyrene box) to 475 μl of 1% PBS-BSA-T and mix        thoroughly—labelled ‘125’.    -   250 μl of 1% PBS-BSA-T is added to 7 further epindorfs which are        labelled 62.5, 31.25, 15.63, 7.61, 3.9, 1.95 and 0.98.    -   250 μl is taken from the 1st epindorf (labelled 125) and        transferred to the next (labelled 62.5). This is mixed        thoroughly.    -   Using a new pipette tip, 250 μl is removed from epindorf        labelled 62.5 and transferred to 31.25, this procedure is        continued down to the 0.98 concentration (125, 62.5, 31.25,        15.63, 7.61, 3.9, 1.95, 0.98)    -   In total 475+(250×7)=2.3 ml: 0.023 g of BSA added to 2.3 ml of        PBS-T.-   11. Add 100 μl aliquots of the allergen sample to the plate along    with the standard allergen samples for the reference curve in    duplicate. The standards usually go in the first two columns on the    left hand side, with the least concentrated on top. Incubate for 1    hour.-   12. Follow stages 5 to 6, completing a total of 5 washes.-   13. Pour 11 ml of 1% BSA-PBS-T (0.11 g of BSA to 11 ml of PBS-T) to    the dispensing tray. Add 11 μl of the biotinylated monoclonal    antibody (refrigerator) and mix thoroughly.-   14. Pipette 100 μl into each well and incubate for 1 hour at room    temperature.-   15. Empty plate and wash as described in stage 12. (5 washes).-   16. Add 11 μl of Streptavidin (freezer) to 11 ml of 1% BSA-PBS-T.    Pipette 100 μl into each well and incubate for 30 minutes. Reserve    any remaining solution in a vial.-   17. Empty plate and wash as described in stage 12 (5 washes).-   18. Make a solution of OPD, by putting the two tablets (in silver    and gold foil) into 20 ml of distilled water (in a glass vial).    Shake quite vigorously in the dark until the tablets have dissolved    (Wrap the vial up either in tin foil or paper towel).-   19. Add a small amount to the remaining solution from stage 16. Wait    for a colour change (positive reaction). Add 200 μl to each well and    incubate for a minimum of 30 minutes in the dark.-   20. Read the plate at 450 nm/405 nm if filter not available.

Examples 1 to 26

The deactivants, as set out in the following table, were used againstDer-f allergens according to the above procedure and the results are asgiven below. Tannic acid was used as a comparator. What was measuredafter treatment with deactivant and tannic acid was the amount ofallergen remaining active after treatment. The ratio of amount ofremaining active allergen after treatment with deactivant and tannicacid is also given.

TABLE Amount of Ratio of remaining Allergen active allergen Amount ofAllergen remaining active after remaining active after after tannic acidDeactivant/Tannic Example Deactivant deactivant treatment treatment AcidTreatment Number 1 Urea 3750 1500 2.500 xxi 2 Polymeric dialdehyde 1325550 2.409 xx 3 Cedarwood oil 1800 750 2.400 i 4 Cyclodextrin 3850 17002.265 xxii 5 hexadecyltrimethylammonium chloride 4075 1800 2.264 ii 6Aluminium chlorohydrate 1675 750 2.233 iii 7 1-propoxy-propanol-2 39501800 2.194 iv 8 Silica Gel (Kent) 2037.5 933.5 2.183 vi 9polyquaternium-10 (Polymer JR-125) 4335 2000 2.168 v 10 Hydrogenated HopOil 1100 550 2.000 xxiii 11 Propylene glycol alginate 3175 1700 1.868vii 12 Poly vinyl pyrrolidone 2450 1425 1.719 xxiv 13 Ammonium sulphate2750 1700 1.618 viii 14 Hinokitol (0.5%) 3065 2000 1.533 ix 15 N-methylpyrrolidone 1600 1175 1.362 xxv 16 L-Ascorbic Acid 2000 1500 1.333 x 17Immobilised Tannic Acid 1550 1175 1.319 xi 18 Aerosol OT 1525 1175 1.298xviii 19 Chlorohexidine 1412.5 1425 0.991 xii 20 Parsley Camphor 12251387.5 0.883 xix 21 Maleic anhydride 1312.5 1500 0.875 xiii 22Anthraquinone sodium salt 1530 2000 0.765 xxvi 23 Hinoki oil 1025 1387.50.739 xiv 24 Composite of AgCl and TiO₂ 1025 1425 0.719 xv 25 Germall II950 1387.5 0.685 xvi 26 Thymol 725 1387.5 0.523 xvii

Examples 27 to 47

The deactivants, as set out in the following table, were used againstDer-p allergens according to the above procedure and the results are asgiven below. What was measured were the amount of allergens remainingafter treatment with deactivant and the amount of allergens remainingafter vacuuming with no deactivant treatment.

TABLE Amount of active Allergen remaining after deactivant ExampleDeactivant treatment Deactivant Amount of active Allergen remainingafter no deactivant treatment but only vaccuming 1 Glutaraldehyde 8163375 xxviii 2 Polymeric dialdehyde 2792 3375 xx 3 Cedarwood oil 33756000 i 4 hexadecyltrimethyl- 2863 4992 ii ammonium chloride 5 Aluminiumchlorohydrate 978 4992 iii 6 1-propoxy-propanol-2 1233 4992 iv 7 SilicaGel (Kent) 1540 4992 vi 8 polyquaternium-10 5463 6250 v (Polymer JR-125)9 Propylene glycol alginate 3781 6250 vii 10 Ammonium sulphate 2325 6250viii 11 Potassium thioglycolate 3092 3375 xxvii Amount of Allergenremaining after no deactivant treatment 12 Hinokitol (0.5%) 2058 3375 ix13 L-Ascorbic Acid 1438 5642 x 14 Immobilised Tannic Acid 1125 5642 xi15 Aerosol OT 4494 5642 xviii 16 Chlorohexidine 2281 4450 xii 17 ParsleyCamphor 2581 4450 xix 18 Maleic anhydride 783 4450 xiii 19 Hinoki oil1644 3400 xiv 20 Composite of 1632 3400 xv AgCl and TiO₂ 21 Thymol 15003400 xvii

Examples 48-51

The following formulations can be made up as carrier compositions foruse in an aerosol for deactivating Der-f and Der-p allergens.

Example 48

Raw Ingredient Description By Weight Item Classification % AnhydrousEthanol (SD Solvent 79.646 Alcohol 40) Alkyl dimethyl benzyl CationicSurfactant 0.106 ammonium saccharinate Corrosion Inhibitor (I) 0.192Corrosion Inhibitor (II) 0.192 Corrosion Inhibitor (III) 0.096 DeionizedWater Water/Solvent 15.768 Carbon Dioxide Propellant 4.000 TOTAL 100.000

Example 49

Raw Ingredient Description by Weight Item Classification % AnhydrousEthanol (SD Solvent *57.000 Alcohol 40) Fragrance #17 Fragrance 0.0500Dow Corning 193 Surfactant 0.025 Surfactant Corrosion Inhibitor (I)0.100 Corrosion Inhibitor (II) 0.100 Deionized Water Water/solvent*14.725 NP-40/Butane 40 Hydrocarbon 28.000 propellant TOTAL 100.000 *=May replace with 95% Ethanol (SD Alcohol 40) at 61.755% by weight and9.970% by weight Deionized water

Example 50

Raw Ingredient Description by Weight Item Classification % AnhydrousEthanol (SD Solvent 79.646 Alcohol 40) Benzyl Benzoate - an Active/ester4.600 acaricide Alkyl dimethyl benzyl Cationic Surfactant 0.106 ammoniumsaccharinate Corrosion Inhibitor (I) 0.192 Corrosion Inhibitor (II)0.192 Corrosion Inhibitor (III) 0.096 Deionized Water Water/solvent11.168 Carbon Dioxide Propellant 4.000 TOTAL 100.000

Example 51

Raw Ingredient Description by weight Item Classification % AnhydrousEthanol (SD Solvent *57.000 Alcohol 40) Benzyl Benzoate Active/ester4.600 Fragrance #17 Fragrance 0.0500 Dow Corning 193 Surfactant 0.025Surfactant Corrosion Inhibitor (I) 0.100 Corrosion Inhibitor (II) 0.100Deionized Water Water/solvent *10.125 NP-40/Butane 40 Hydrocarbon 28.000propellant TOTAL 100.000 *= May replace 95% Ethanol (SD Alcohol 40) at61.755% by weight and 5.370% by weight Deionized water.

1. The method for deactivating a Der-f and/or a Der-p allergen whichcomprises spraying the allergen with a deactivating effective amount ofa liquid composition comprising xi) immobilised tannic acid.
 2. Themethod according to claim 1 in which the composition is an aqueouscomposition.
 3. The method according to claim 1 in which the compositionadditionally comprises one or more of a fragrance, a surfactant, anantimicrobial agent, a corrosion inhibitor and a miticide.